Single cell gene expression for diagnosis, prognosis and identification of drug targets

Abstract

Methods are provided for diagnosis and prognosis of disease by analyzing expression of a set of genes obtained from single cell analysis. Classification allows optimization of treatment, and determination of whether on whether to proceed with a specific therapy, and how to optimize dose, choice of treatment, and the like. Single cell analysis also provides for the identification and development of therapies which target mutations and/or pathways in disease-state cells.

1 - 12 . (canceled) 
     
     
         13 . A method of analyzing a heterogeneous tumor biopsy from a subject, comprising:
 randomly partitioning cells from the biopsy into discrete locations;   performing transcriptome analysis on at least 50 genes of the individually partitioned cells; and   using transcriptome data to identify one or more characteristic of said tumor.   
     
     
         14 . The method of  claim 13 , wherein the performing step is done without prior enrichment of a cell type. 
     
     
         15 . The method of  claim 13 , wherein the characteristic is the presence, absence, or number of cancer cells. 
     
     
         16 . The method of  claim 13 , wherein the characteristic is the presence, absence or number of stem cells, early progenitor cells, initial differentiated progenitor cells, late differentiated progenitor cells, or mature cells. 
     
     
         17 . The method of  claim 13 , wherein the characteristic is effectiveness of a therapeutic agent in eliminating one or more of the cells. 
     
     
         18 . The method of  claim 13 , further comprising using said characteristic to diagnose said subject with cancer or a cancer stage. 
     
     
         19 . The method of  claim 13 , wherein the characteristic is activity of a signaling pathway. 
     
     
         20 . The method of  claim 19 , wherein said signaling pathway is specific to a cancer stem cell, a differentiated cancer cell, a mature cancer cell, or combination thereof. 
     
     
         21 . A method of identifying a signaling pathway utilized by a disease-state cell, comprising:
 randomly partitioning cells from a heterogeneous sample;   performing transcriptome analysis on said partitioned cells;   using transcriptome analysis to identify at least one disease-state cell;   comparing said transcriptome analysis of said at least one disease-state cell to transcriptome of:
 a) a non-disease state cell; 
 b) a different disease-state cell; and 
 c) a disease-state stem cell; and 
   identifying a signaling pathway that is expressed in (i) the disease-state cell, (ii) the disease-state stem cell, and (iii) optionally in the different disease-state cell, but not in a non-disease-state cell, thereby identifying a signaling pathway utilized by a disease-state cell.   
     
     
         22 . The method of  claim 21 , wherein said disease state is cancer, ulcerative colitis or inflammatory bowel disease. 
     
     
         23 . The method of  claim 21 , wherein said signaling pathway is required for survival of said disease state cell. 
     
     
         24 . A method for diagnosing a subject with a condition comprising,
 randomly partitioning cells from a heterogeneous sample;   performing a first transcriptome analysis on partitioned cells;   using transcriptome analysis to identify at least one disease-state cell by comparing said first transcriptome analysis from said at least one disease-state cell to a second transcriptome analysis from a non-disease state cell, thereby diagnosing the presence or absence of a condition associated with said disease state cell in said subject.   
     
     
         25 . The method of  claim 24 , wherein said disease state is breast cancer, colon cancer, ulcerative colitis or inflammatory bowel disease. 
     
     
         26 . The method of  claim 21 , wherein said transcriptome analysis comprises analyzing expressed RNA, non-expressed RNA, or both. 
     
     
         27 . The method of  claim 21 , wherein said transcriptome analysis is whole transcriptome analysis. 
     
     
         28 . The method of  claim 21 , wherein said disease state cell is a breast cancer stem cell. 
     
     
         29 . A method for screening for a therapeutic agent comprising:
 exposing a first subject with disease-state cells to one or more test agents;   obtaining a heterogeneous tumor biopsy from the subject from a region of interest;   performing transcriptome analysis on at least one individual cell from said heterogeneous tumor biopsy,   wherein said biopsy comprises one or more disease state cells; and   comparing said transcriptome analysis to a transcriptome derived from either:
 (i) a second subject without the disease-state cells; or 
 (ii) the first subject prior to said exposing step; and 
   identifying an agent that affect transcriptome of cells from the test area to be more like those of the second subject or the first subject prior to exposure.   
     
     
         30 . A method of determining the potential effectiveness of a therapeutic agent against a disease, comprising:
 separating a first population of disease-state cells into individual locations, wherein said individual locations comprise an individual cell;   determining the expression level of at least one nucleic acid or protein from at least one of said individual cells, thereby producing a disease-state expression signature;   exposing a second population of disease state cells to an agent;   separating said second population of disease-state cells into individual locations, wherein said individual locations comprise an individual cell;   determining the expression level of at least one nucleic acid or protein from at least one of said individual cells from said second population; and   comparing said expression level from said individual cell from said second population to said disease-state expression signature, thereby determining the effectiveness of said agent against said disease.   
     
     
         31 . The method of  claim 30 , wherein said exposing step is performed in vivo, 
     
     
         32 . The method of  claim 30 , wherein said first population and said second population are isolated from a subject. 
     
     
         33 . The method of  claim 32 , wherein said subject s a human. 
     
     
         34 . The method of  claim 30 , wherein said disease is cancer, ulcerative colitis or inflammatory bowel disease. 
     
     
         35 . The method of  claim 30 , wherein said nucleic acid or said protein is a cancer stem cell marker. 
     
     
         36 . The method of  claim 30 , wherein said expression level is mRNA expression 
     
     
         37 . The method of  claim 33 , wherein determining said mRNA expression level comprises detection of expression or lack of expression of 10 or more nucleic acids. 
     
     
         38 . The method of  claim 30 , wherein said expression level is protein expression level.) 
     
     
         39 . The method of  claim 30 , wherein said separating steps comprise exposing said population of cells to an antibody that specifically binds a protein present on said individual 
     
     
         40 . A method of determining likelihood of a response by a subject to a therapeutic agent, comprising:
 separating a population of cells from a subject into individual locations, wherein said individual locations comprise an individual cell and wherein at least one of said individual cells is a disease-state cell;   determining the expression level of at least one nucleic acid or protein from at least one of said disease-state individual cells, wherein said nucleic acid or protein is a target of a therapeutic agent; and   determining likelihood of a response by a subject based on the expression level of said at least one nucleic acid or protein.   
     
     
         41 . The method of  claim 40 , wherein said expression level is mRNA expression level. 
     
     
         42 . The method of  claim 41 , wherein determining said mRNA expression level comprises detection of expression or lack of expression of 10 or more nucleic acids. 
     
     
         43 . The method of  claim 42 , wherein said expression level is protein expression level. 
     
     
         44 . The method of  claim 42 , wherein said separating step comprises exposing said population of cells to an antibody that specifically binds a protein present on said individual cells. 
     
     
         45 . The method of  claim 40 , wherein said therapeutic agent is an anti-cancer agent. 
     
     
         46 . The method of  claim 22 , wherein said disease state is an epithelial-derived cancer. 
     
     
         47 . The method of  claim 22 , wherein said disease state is breast cancer or colon cancer.